The nature, development, interrelationship and functional significance of the several subclasses of muscarinic acetylcholine receptor (mAChR) will be studied in rat brain. Multiple receptor populations (with respect to both agonist and antagonist binding) and a regional heterogeneity of mAChR have been demonstrated. In the proposed research, the biochemistry and pharmacology of mAChR in the different receptor subpopulations and from different brain areas will be studied using radiolabelled ligand binding techniques. This study will include a consideration of agonist and antagonist binding affinity, kinetics and thermodynamics. A comparison of binding properties with two receptor-regulated postsynaptic processes, cGMP synthesis and phospholipid metabolism, is expected to reveal the functional significance of the multiple mAChR populations and regional heterogeneity. Determinations of the molecular size of mAChR in different states or from different brain regions will indicate possible differences in the coupling of mAChR to postsynaptic effector mechanisms and/or regulatory subunits. Modification of mAChR binding properties and postsynaptic responses, as well as ACh metabolism, due to prolonged exposure to receptor ligands, guanine nucleotides or heavy metals or treatment with sulfhydryl reagents will be studied in vitro using brain minces. The importance of the hydrophobic membrane environment in the control and maintenance of receptor function will be investigated in experiments in which membrane compostition and/or structure is altered through enzymatic treatment, freeze-thaw induced fusion with exogenous liposomes, and controlled membrane disaggregation by sonication in the presence of small amounts of chaotropic ions or detergents. A separation of the receptor from (and a reconstitution of the receptor with) the subunit with which GTP interacts to influence receptor binding will be attempted using affinity chromatographic techniques. The proposed research is expected to provide information on the regulation of receptor function which may play an important role in neuronal synaptic plasticity. In addition, information will be obtained concerning possible biochemical bases of neuronal dysfunction as well as the mechanisms underlying drug action in the central nervous system.